Biology subjects

Roy, S.

Publications and source records attributed to Roy, S..

At least 55 records · Page 3Linked to original sources

bioRxiv2026

Soluble LAG-3 Identifies a Dynamic Early T Cell Activation Window in self-reactivity, Type 1 Diabetes, and Broader Immune Responses.

Aims/hypothesisType 1 diabetes is a complex autoimmune disorder in which autoreactive CD4 and CD8 T cells destroy pancreatic beta-cells, resulting in insulin deficiency and hyperglycemia. Although genetic susceptibility, particularly certain HLA alleles, contributes to disease risk, not all genetically predisposed individuals develop Type 1 diabetes. Screening first degree relatives (FDRs) for islet autoantibodies (GAD65, IAA, IA-2, ZnT8) helps detect autoimmune activity. However, these serum markers arise only after T-helper cell activation, limiting early intervention opportunities. Since protein antigen recognition by B cells requires T-helper cell assistance through linked recognition, T cell activation precedes B cell activation and autoantibody production. Activation of these T cells leads to shedding of the immune-regulatory (activation) surface protein LAG-3 (Lymphocyte Activation Gene-3 or CD223), generating its soluble form, sLAG-3, that is detectable in circulation. We hypothesized that sLAG-3 may serve as an early biomarker of autoimmune activity preceding islet autoantibody development in type 1 diabetes. MethodsPlasma sLAG-3 levels were measured longitudinally in female diabetes-prone NOD mice and analyzed in relation to islet antigen-specific CD4 T cell expansion and diabetes onset. To mechanistically link autoreactive T cell activation to sLAG-3 release. Naive autoreactive C6.6.9 TCR-transgenic (TCR-Tg) CD4 T cells were adoptively transferred into NOD.SCID mice and longitudinal assessment for plasma sLAG-3, beta-cell antigen specific CD4 T cell tetramer profiles, and circulating insulin (Ins2) mRNA to determine ongoing beta-cell stress. In parallel, sLAG-3 levels were analyzed from different human cohorts, including FDRs of individuals with type 1 diabetes, using cross-sectional and longitudinal approaches. ResultsIn murine models, elevated sLAG-3 correlated with expansion of islet-specific CD4 T cells that preceded hyperglycemia and diabetes onset. In the adoptive transfer model, early increases in sLAG-3 and circulating Ins2 mRNA marked immune activation and emerging beta-cell stress prior to overt diabetes. In our human cohorts, sLAG-3 was detectable in autoantibody-negative and single-autoantibody-positive FDRs, with higher levels observed in progressors compared to non-progressors, and associated with high-risk HLA genotypes. Conclusions/interpretationThese findings identify sLAG-3 as a candidate biomarker of early T cell activation in type 1 diabetes that may precede islet autoantibody development. Integration of sLAG-3 with antigen-specific T cell and beta-cell stress markers could improve early risk stratification and inform preventive strategies before substantial loss of beta-cell. Prospective longitudinal studies aligned to seroconversion are required to validate sLAG-3 as a surrogate marker of early disease activity. Research in contextO_ST_ABSWhat is already known about this subject?C_ST_ABSO_LIBefore the clinical onset of hyperglycemia, type 1 diabetes is characterized by a prolonged preclinical phase in which autoreactive B and T cells mediate progressive beta-cell destruction. C_LIO_LICurrent risk stratification strategies rely mainly on genetic susceptibility (genomic DNA) and the detection of islet autoantibodies in plasma/serum. C_LIO_LIIslet autoantibodies arise only after CD4 T cell activation and therefore do not capture the earliest stages of immune dysregulation. C_LIO_LIConsequently, biomarkers that directly reflect early pathogenic T cell activity prior to, or independent of, seroconversion remain limited and insufficiently validated. C_LI What is the key question?Can plasma sLAG-3 levels, beta-cell antigen-specific CD4 T cell tetramer expression, and circulating Ins2 mRNA serve as very early biomarkers of autoimmune activity in type 1 diabetes and serve to better inform risk stratification, thereby informing preventive intervention strategies for the clinician? What are the new findings?O_LIsLAG-3 increases transiently during early antigen-specific CD4 T cell activation stage, precedes hyperglycemia in mouse models, and is elevated in autoantibody-negative and single-autoantibody-positive first-degree relatives who later progress to type 1 diabetes. C_LIO_LIsLAG-3 was associated with beta-cell antigen-specific CD4 T cell expansion, assessment of stress induced beta cell Ins2 mRNA release and high-risk HLA genotypes, indicating early autoimmune activation rather than established disease. C_LI How might this impact clinical practice in the foreseeable future?These findings support sLAG-3 as a candidate early biomarker of T cell activation, before or at the earliest stages of islet autoantibody development in some at-risk individuals. Integration of plasma sLAG-3 with beta-cell antigen specific CD4 T cell profiling and insulin mRNA measurements could complement current autoantibody-based screening, improve risk stratification, and enable earlier preventive interventions to preserve beta-cell function in patients at-risk for type 1 diabetes.

immunology↗

bioRxiv2026

A Multiplex Droplet Digital PCR Assay for Chromosome Copy Number Determination in Candida albicans

Chromosome copy number variation (CNV) is a major contributor to genome plasticity and adaptation in Candida albicans, a leading fungal pathogen of humans. Aneuploidy, defined as deviations from the normal diploid chromosome set, rapidly alters gene dosage, enabling tolerance to host-imposed and antifungal stress. Accurate detection and quantification of chromosomal copy number changes are thus essential to dissect the mechanisms by which C. albicans adapts and evolves. Here, we describe the development, optimization, and validation of a six-color, 16-plex droplet digital PCR assay for simultaneous quantification of all C. albicans chromosome arms in a single reaction. Each target is detected by a unique dual-color or single-color combination of probes, enabling high-order multiplexing through binary fluorescence encoding. Following optimization of probe concentrations, PCR cycling parameters, genomic DNA extraction and pre-treatment with restriction enzymes, the assay provides accurate, reproducible chromosome-level copy number estimates that correlate closely with WGS results across euploid and aneuploid isolates. Compared to whole-genome sequencing, the assay is rapid, cost-effective, and scalable, requiring minimal DNA input and allowing high-throughput analysis of large isolate collections. The 16-plex assay thus provides a platform for dissecting genome instability and adaptive evolution in C. albicans. Article SummaryWe developed and validated a 16-plex droplet digital PCR assay that estimates chromosome dosage across the entire genome of the human fungal pathogen C. albicans in a single reaction. The assay uses six fluorescent colors and unique color combinations to track one marker on each chromosome arm, enabling rapid detection of aneuploidy (extra or missing chromosomes). Results closely matched whole-genome sequencing for isolates with simple aneuploid forms and detected low-frequency trisomic clones in mixed populations. With optimized DNA preparation, this method provides a practical tool for screening genome instability in research and clinical settings.

microbiology↗

bioRxiv2026

Mechanism of O2 Activation for Conversion of Fatty Acids into Terminal Alkenes in a Membrane-bound Metalloenzyme UndB

Multinuclear metal centers catalyze some of the most challenging oxidative transformations in biological and chemical catalysis, yet the molecular principles controlling oxygen activation in non-heme diiron enzymes remain unclear. Structural studies of desaturase-like diiron enzymes frequently reveal elongated Fe-Fe separations lacking canonical bridging motifs, whereas spectroscopic measurements indicate substantial metal-metal coupling during catalysis, creating a longstanding mechanistic paradox between structural and spectroscopic observations. Here we show that oxygen activation can be dynamically regulated by fluctuations in metal-metal distance through extensive multiscale simulations of the membrane-bound fatty-acid decarboxylase UndB. UndB catalyses the conversion of naturally abundant free fatty acids into terminal 1-alkenes-valuable hydrocarbon precursors for sustainable biofuels, and exhibits unusual hydrogen peroxide formation that has led to conflicting mechanistic proposals. Combining quantum mechanics/molecular mechanics (QM/MM) simulations with free energy analysis and quantum chemical calculations, we identify a catalytically competent ensemble in which transient Fe-Fe contraction establishes effective metal-metal coupling and enables formation of a peroxodiiron(III/III) intermediate despite elongated resting-state structures and the absence of canonical bridging motifs. The computed free-energy landscape quantitatively reproduces experimentally inferred catalytic barriers and accounts for the substantial H2O2 formation observed during catalysis. The reactive state resembles a P-like intermediate rather than the canonical diamond-core Q species commonly invoked for related diiron enzymes. Comparison with magnetic coupling and mechanistic features reported for related non-heme diiron enzymes, including Alkane monooxygenase AlkB, Stearoyl-CoA Desaturase 1 (SCD1) and the soluble decarboxylase UndA, suggests that dynamic metal-metal distance modulation represents a general mechanism governing oxygen activation across diverse diiron enzyme families. These findings establish metal-metal distance modulation as a previously unrecognized control parameter for oxygen activation in diiron and related multinuclear metalloenzymes, reconciling structural and spectroscopic observations and revealing how protein conformational dynamics regulate electronic coupling to control oxidative catalysis, thereby suggesting a general principle for tuning reactivity in multinuclear catalysts.

biophysics↗

bioRxiv2026

Dual Effect of RNA-like Polyelectrolytes on Stability and Dynamics of Biomolecular Condensates: A Tale of Competitive Short and Long-range Interactions

Heterotypic biomolecular condensates underpin the spatiotemporal organization of cellular components and enable precise biological regulation. These condensates frequently comprise proteins and RNA at variable stoichiometries, with RNA acting as a key modulator through its ability to engage in both long-range electrostatic and short-range specific interactions. How such RNA-like components regulate condensate stability and dynamics across distinct interaction regimes, however, remains unclear. Here, we examine the role of RNA-like polyelectrolytes in tuning the stability and material properties of heterotypic condensates formed by designed short peptide sequences spanning a continuum from long-range electrostatics-to short-range hydrophobic interactions. By systematically strengthening short-range hydrophobic interactions while varying polyelectrolyte composition, we uncover a dual, regime-dependent role of RNA-like species. In electrostatics-dominated condensates, excessive polyelectrolyte concentration rapidly destabilizes droplets due to enhanced long-range repulsive interactions while the droplets retain stability until intermediate polyelectrolyte concentration fairly well. In contrast, in strongly hydrophobic condensates, polyelectrolytes function as multivalent sticker hubs, stabilizing condensates through favourable peptide- polyelectrolyte interactions. Polyelectrolyte enrichment within condensates is nonlinear with respect to mixing fraction and saturates at largest cluster mole fractions of [~]0.2-0.25. Condensate dynamics reflect this interplay: polyelectrolyte diffusivity is 20-40% higher in electrostatics dominated systems than in hydrophobic ones, while in extremely hydrophobic condensates, polyelectrolytes diffuse [~]50% more slowly than peptides, indicative of scaffold-like behaviour. Together, these results reveal tuneable and opposing roles of RNA-like polyelectrolytes in shaping condensate stability, dynamics, and morphology across diverse interaction regimes.

biophysics↗

bioRxiv2026

DNA-templated Chiral Metamaterial Array as Information Bits

The growth of digital information demands physically secure information bits that combine intrinsic randomness with multi-layered optical readout. Digital metamaterials with physical unclonable function characteristics are promising, but nearly all examples operate at microwave or terahertz frequencies. Extending digital metamaterials to the visible range requires a physical property that allows binary (0/1) encoding via nanoscale geometry and is orthogonal to intensity-or color-based imaging. Chiral plasmonic metamaterials satisfy these criteria perfectly: their broken mirror symmetry yields chirality polarity in the visible spectrum whose sign and magnitude directly encode bit values ("0", spin-up; "1", spin-down), while remaining invisible under linear polarization. Here, we realize visible-range digital metamaterials by programming self-assembled DNA origami templates with asymmetrically placed gold nanorods to create discrete 3D chiral metamolecules. Using on-surface DNA origami assembly and the Silica Microsphere-Assisted Patterning by Liquid Elimination (SiMPLE) method, we fabricate large-scale bit arrays on optical active glass substrate in solution with [~]1.33 m lattice spacing, [~]86% site occupancy, and [~]12-month stability after silicification. These bottom-up fabricated digital metamaterial array exhibits two independent security layers: (1) a macroscopic spatial pattern only visible by dark-field microscopy, and (2) hidden binary information stored in the single-particle chiroptical response, read out by position-resolved circular dichroism spectroscopy. Quantitative analysis confirms reliable bit encoding and high optical randomness arising from slight structural variations. By leveraging the polarity of plasmonic chirality to translate molecular-scale handedness into robust visible-range digital signals, this work establishes a scalable nanophotonic platform for secure optical information storage, authentication, and encryption.

bioengineering↗

bioRxiv2026

Conformalized Multiview Learning

SO_SCPLOWUMMARYC_SCPLOWThere is an overwhelmingly large body of literature and numerous algorithms already available on "multimodal AI", based on different modeling and fusion paradigms. As multimodal AI becomes fully integrated into healthcare applications, the ability to say "Im not sure" or "I dont know" when uncertain is a necessary capability for safe clinical deployment, requiring the augmentation of current multimodal AI techniques. Our goal in this work is not to add yet another method to the growing toolbox of multimodal AI, but rather to (i) clarify how existing supervised multimodal AI methods can be understood through the lens of uncertainty quantification, and (ii) develop a unified framework that provides valid and interpretable uncertainty estimates for multimodal AI practitioners. We introduce Coracle, a conformalized framework for multimodal AI that seamlessly adapts to early, late, and intermediate fusion. Coracle provides theoretical marginal confidence guarantees and achieves valid finite-sample coverage without relying on distributional assumptions. Through extensive simulations and analyses of diverse biomedical multiview datasets, we demonstrate that Coracle consistently outperforms existing non-conformalized methods in both uncertainty estimation and calibration. By equipping published multiview models with valid, interpretable confidence measures, Coracle advances the development of trustworthy, uncertainty-aware statistical tools for biomedical decision support. Coracle is publicly available as an open-source R package at https://github.com/himelmallick/Coracle.

systems biology↗

bioRxiv2026

Quantitative Deciphering of Mammalian Histamine Receptors through Mathematical Genomics

Histamine receptors (HRH1-HRH4) are G-protein coupled receptors (GPCRs) that mediate essential physiological functions, including neurotransmission, gastric acid secretion, immune regulation, and presynaptic autoregulation. Despite their importance, systematic comparative analyses across mammalian histamine receptor sequences remain scarce. In this study, we performed a comprehensive evaluation of HRH1-HRH4 across multiple mammalian species, integrating sequence homology, invariant residue mapping, amino acid substitutions, compositional frequencies, Shannon entropy, polystring distributions, intrinsic disorder profiling, and phylogenetic clustering. HRH2 and HRH1 were highly conserved among primates, HRH3 showed strong cohesion within rodents, while HRH4 exhibited pronounced divergence consistent with immune-related specialization. Invariant residues localized to transmembrane helices and activation motifs (D107, W428/Y431, NPxxY), underscoring strict evolutionary constraints on ligand binding, receptor stability, and G-protein coupling. Substitutions were confined to non-essential lipid-facing and loop regions, predominantly conservative in nature, enabling diversification without disrupting the GPCR fold. Amino acid frequency and entropy analyses revealed hydrophobic dominance with subtype-specific enrichment of polar residues, while disorder profiling identified HRH1 as the most dynamic and HRH2 as the most structurally constrained. Polystring analysis highlighted conserved motifs (WWW, PP) alongside subtype- and species-specific repeats, reflecting evolutionary strategies balancing receptor stability with adaptive flexibility. Phylogenetic clustering confirmed subtype-specific cohesion, with HRH3 and HRH4 forming compact clades, HRH1 showing moderate dispersion, and HRH2 forming the most isolated cluster. Collectively, these findings demonstrate that mammalian histamine receptor evolution is governed by conserved biophysical cores and selective variability, offering insights into structural conservation, functional diversification, and translational relevance for drug design and model selection.

bioinformatics↗

bioRxiv2026

Stress-Survival Pathway Profiling Reveals MCM10 as a Candidate Biomarker of Hispanic Colorectal Cancer Disparities

BackgroundColorectal cancer (CRC) remains the second leading cause of cancer mortality in the U.S., with significant racial and ethnic disparities in incidence, survival, and mortality rates. The rising incidence of early-onset CRC and the frequent presentation at advanced stages of CRC among the Hispanics makes this an important ethnic group to study. A deeper understanding of the complex interplay between molecular, genetic, and environmental factors is critical for developing targeted therapies to reduce the prevalence in specific racial and ethnic groups, such as Hispanics. This study explores the role of stress-survival pathway (SSP) genes in early-onset and late-stage CRC, primarily focusing on disparities between Hispanics and Non-Hispanic Whites (NHWs). Additionally, this study investigates the role of MCM10, in contributing to CRC disparities among the Hispanic populations with an emphasis of early-onset and late-stage CRC Hispanics. MethodsOne of our previous studies had identified some SSP protein coding genes associated with CRC. The transcript and protein level expressions of these genes were validated in CRC cell lines, cDNA arrays, and tissue microarrays, using qRT-PCR and immunohistochemistry, respectively. The transcript level expressions of differentially expressed SSP genes were further evaluated in Hispanic and NHWs tumor tissues, including early onset and late-stage cohorts. Additionally, we performed cellular, physiological, and functional assays to explore the role of MCM10 in tumor progression, before and after siRNA mediated knockdown of MCM10. High-throughput transcriptomic and proteomic analyses were performed to reveal the underlying molecular mechanism. ResultsWe observed the differential expressions of twelve SSP genes in CRC cell lines, at the transcript level; that of ten genes in early and late stages using cDNA arrays, and nine genes at the protein levels using tissue microarrays and immunohistochemistry. Hispanic CRC samples showed differential expression of all nine SSP genes compared to NHWs, in early-onset, and late-stage CRC, with all genes being upregulated other than CDK4 and PRDX4 in late-stage CRC and CDK4 in early-onset CRC. Our functional study demonstrated that MCM10 knockdown in CRC cell lines significantly reduced cell proliferation, growth, invasion, and migration by inducing cell cycle arrest, apoptosis, and reactive oxygen species pathways. The integrative RNA-sequencing and proteomics study identified that PPFIA1 could be associated with MCM10 mediated cancer progression. The role of MCM10 and PPFIA1 in cancer progression has also been validated by CRISPR-Cas9 mediated MCM10 knock out, using a Hispanic CRC patient derived organoid. ConclusionThe differential expression of SSP genes suggest a potential molecular contribution to CRC disparities in the Hispanic population. The essential oncogenic role of MCM10 and its axis with PPFIA1 was identified; this could lead to a new avenue for therapeutics targeting the MCM10-PPFIA1 axis, thereby, combating early-onset and advanced-stage CRC in this high-risk ethnic group. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=177 HEIGHT=200 SRC="FIGDIR/small/698057v2_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@11528c8org.highwire.dtl.DTLVardef@288d15org.highwire.dtl.DTLVardef@466703org.highwire.dtl.DTLVardef@150890_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

Thioredoxin interacting protein (TXNIP), a redox regulator, mediates the RAPGEF3/4 signaling dependency in primary melanoma

Progression of cutaneous primary melanoma that arises from melanocytes leads to lethal metastatic disease. Molecular mechanisms that control the growth of primary melanoma in the skin and promote progression are not fully understood. Previously we showed that RAP guanine exchange factors EPAC1/2 (Exchange Protein Activated by cyclic AMP) promote the growth of primary melanoma and loss of dependency on EPACs is associated with metastatic progression. In this study, we show that EPACs are activated during malignant transformation of melanocytes, and chemical inhibition or genetic deletion of EPAC inhibits melanomagenesis in Braf/Pten mice. Low expression of EPAC mRNA and its effector RAP1-GTP protein in primary melanoma correlate with better recurrence-free survival. RNAseq analysis of matched primary and metastatic melanoma cells treated with an EPAC inhibitor showed that TXNIP, an important regulator of redox homeostasis, is a downstream effector of EPAC signaling. We also show that EPACs promote melanoma growth by regulating redox homeostasis and mitochondrial ROS through activation of mechanistic target of rapamycin complex 1 (mTORC1) that stabilizes hypoxia-inducible factor 1-alpha (HIF-1), a transcriptional activator of redox regulator TXNIP and glycolytic enzymes. Our data suggest that targeting mechanisms that melanoma cells employ to bypass EPAC dependency is a potential therapeutic approach. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=134 SRC="FIGDIR/small/696903v2_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1967afaorg.highwire.dtl.DTLVardef@63b907org.highwire.dtl.DTLVardef@1089da4org.highwire.dtl.DTLVardef@a5b2ef_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

Conformational and Environmental Determinants of RNA Solvation Dynamics: Roles of Intrinsic Flexibility, Allostery, and Protein Binding

Solvation dynamics play a central role in shaping nucleic acid structure, flexibility, and recognition, yet their molecular origins remain poorly understood for RNA, whose diverse architectures and intrinsic conformational plasticity far exceed those of DNA. Here, we present the atomistic, microsecond-scale computational dissection of solvation dynamics in two structurally homologous but functionally distinct viral RNAs--BIV TAR and HIV-2 TAR--in both apo and peptide-bound states mimicking the salt concentration of experimental time-resolved fluorescent spectroscopic measurement. By combining high-temporal-resolution solvation time correlation functions with detailed energy decomposition analyses, we uncover how water, ions, and RNA motions cooperatively shape relaxation across ultrafast to nanosecond timescales. Our results reveal that, unlike DNA, where slow components primarily reflect long-lived hydration and ion condensation, RNA can generate slow solvation decay either through long-lived hydration or through its own internal conformational fluctuations, such as involving spontaneous base-flipping events. Peptide binding modulates this conformational landscape in strikingly system-specific ways: BIV TAR RNA undergoes classical fluctuation quenching, where TAT binding suppresses RNA motions and shifts relaxation toward solvent-RNA compensation, whereas HIV-2 TAR RNA exhibits a non-classical redistribution of solvent-ion-peptide correlations stemming from its weaker and more dynamic binding interface with TAT. The dominant slow decay in HIV-2 apo TAR maps directly onto an allosteric communication channel previously identified from structural analyses, demonstrating that solvation responses can sensitively report on RNA allostery. Together, this study bridges the experimental observations of time resolved fluorescence spectroscopy with mechanistic molecular insight, establishes solvation dynamics as a powerful probe of RNA conformational energetics, and highlights how subtle differences in RNA-protein recognition can imprint distinct signatures on hydration and ion reorganization.

biophysics↗

bioRxiv2025

Behavioral divergence between a generalist and a specialist mosquito despite minimal differentiation across chemosensory receptor gene families

Oviposition site choice is a major determinant of habitat selection in insects, yet its behavioral and genetic bases remain poorly understood. Although model organisms such as Drosophila melanogaster provide well-established systems for studying oviposition, disentangling foraging from egg-laying decisions remains challenging because females use the same substrates for feeding and reproduction. Here, we identify a pair of sister mosquito species that exhibit contrasting oviposition strategies and provide a promising system for investigating how divergence in sensory information processing shapes behavioral shifts between generalist and specialist taxa. Using binary-choice assays, we tested oviposition preferences in 1,046 gravid females from both field and laboratory populations. Anopheles gambiae females were highly selective and deposited nearly all eggs in a single water source, consistent with specialist behavior. In contrast, females of its sibling species, An. coluzzii, adopted a generalist strategy marked by a more even distribution of eggs across available substrates. To identify chemosensory receptor genes that diverge between the cryptic species and may underlie these behavioral differences, we analyzed amino acid substitutions across odorant, ionotropic, and gustatory receptor families using whole-genome sequencing data. Although differentiation was limited at the gene-family level, several individual divergent loci associated with carboxylic acid, amine, and other volatile detection pathways emerged as candidate drivers of species-specific oviposition behavior. These findings suggest that shifts in sensory perception during egg laying may facilitate behavioral and ecological specialization, and that phenotypic divergence in environmental sensing can arise before substantial genome-wide differentiation.

evolutionary biology↗

bioRxiv2025

Emergent simplicities in an individual cell's stochastic response to disruptive change

Here we seek and find recurring patterns of behaviors in the stochastic response of an individual bacterial cell to a disruptive change in growth conditions. Building on the known scaling law that a single timescale, a cellular unit of time, governs stochastic growth and division of individual bacterial cells under constant growth conditions, here we show using experimental data that a dynamic rescaling of the cellular unit of time captures the predominant effect of temporal variations in environmental conditions. Furthermore, we identify the instantaneous exponential growth rate as the scaling factor that scales the internal clocks of the cells to the laboratory time. Our results reveal the natural representation for these time-dependent dynamics. When recast in its terms the cell age distribution for suitable initial conditions evolves under time-invariant rules even as growth conditions remain dynamic! Through the experimental realization at different temperatures of otherwise identical disruptive changes, we not only substantiate the general applicability of the cellular frame of reference but also uncover more emergent simplicities. Motivated by this representation, when time and instantaneous growth rate are expressed in terms of their naturally dimensionless counterparts, remarkably consistent patterns are revealed. These include a unimodal-bimodal-unimodal transition in the shape of the instantaneous growth rate distribution as cells initially in homeostasis experience a disruptive change in nutrient quality and subsequently recover and attain a new homeostasis. The remarkable scaling of the pattern of responses across temperatures suggests that the organizational rules and processes governing the response to disruptive change in nutrient quality remain the same at different temperatures. While the progression of the response appears to proceed at different tempos at different temperatures, upon shifting from the laboratory to the cellular frame of reference these changes progress at the same pace even at different temperatures.

biophysics↗

bioRxiv2025

PARP1 deficiency induces aging-associated cardiac failure via activation of Akt signalling

PARP1, a poly-ADP-ribose transferase, plays a critical role in maintaining genomic stability, transcription, cellular metabolism, and cell death. PARP1 inhibition protects cardiomyocytes against oxidative and genotoxic stress. However, the role of PARP1 in aging-associated heart failure remains poorly explored. In the current study, we report that PARP1 levels are downregulated in aging mouse hearts, and PARP1 deficiency induces aging-related cardiac remodelling and contractile dysfunction in mice. PARP1 deficient mice hearts exhibit spontaneous activation of the Akt signalling pathway, leading to the development of aging-related cardiac hypertrophy and fibrosis. Our findings reveal two distinct mechanisms by which PARP1 regulates Akt signalling, direct interaction of PARP1 with Akt and the transcriptional regulation of phosphatases like PTEN, a negative regulator of Akt signalling. PARP1 binds and inhibits Akt by poly-ADP-ribosylation at E40 and E49 residues, which impairs Akt membrane recruitment and subsequent activation. Inhibition of Akt reversed hypertrophy in PARP1-depleted cardiomyocytes and improved the contractile dysfunction in PARP1-deficient hearts. These findings reveal a previously unrecognized regulatory role for PARP1 in aging-associated cardiac failure.

physiology↗

bioRxiv2025

Neonatal morphine and HIV synergy induce persistent neuroimmune and anxiety-related transcriptional states

BackgroundEarly-life opioid exposure can disrupt neurodevelopment and heighten vulnerability to anxiety and affective disorders, particularly in individuals with HIV. MethodsUsing single-cell RNA sequencing (scRNA-seq), we profiled adolescent brains from wild-type and HIV-1 transgenic (Tg26) mice exposed to morphine during postnatal days 2-7. ResultsMorphine exposure in Tg26 mice resulted in a highly dysregulated microglial phenotype, characterized by the ectopic upregulation of genes encoding neuronpeptides (Avp, Hcrt, and Pmch), while simultaneously showing a reduction in both inflammatory and homeostatic markers (Map3k6, Lgals3, Ccl3). Microglia also showed enhanced expression of dynorphin (Pdyn) and {kappa}-opioid receptor (Oprk1) signaling modules implicated in dysphoria and stress-induced negative effects. Furthermore, transcriptomic mapping revealed cell-type-specific neuronal adaptations: cholinergic neurons upregulated genes linked to anxiety and arousal (Avp, Oxt), GABAergic neurons upregulated genes linked to condition and aversive behavior, whereas glutamatergic neurons enriched for transcripts associated with thigmotaxis and fear behaviors. ConclusionsTogether, these findings demonstrate that brief neonatal morphine exposure in an HIV-inflamed milieu induces persistent, cell-type-specific neuroimmune and neurotransmitter reprogramming that engages the dynorphin-KOR pathway and predisposes to anxiety- and aversion-related behaviors.

neuroscience↗

bioRxiv2025

Distribution of carbon monoxide-oxidizing microorganisms along a chronosequence on Piton De La Fournaise volcano

Volcanic soils provide a unique environment for studying microbial colonization and succession due to their extreme conditions and distinct geochemical profiles. This study focused on carbon monoxide (CO)-oxidizing microbial communities in volcanic soils of varying ages at Piton De La Fournaise (Reunion island). Soil samples from three sites were analyzed to assess microbial community structure using 16S rRNA gene sequencing and metagenomic analysis to identify functional genes involved in CO oxidation. The activity of CO oxidizing microbes in soils was measured. Phylum-level analysis showed increasing Acidobacteriota and Chloroflexota, decreasing Actinomycetota and Bacteroidota, and stable Pseudomonadota, while class-level patterns included rising Alphaproteobacteria and Acidobacteriia, with Ktenobacteria emerging in the oldest soils. CO dehydrogenase-related genes were found in 17 metagenome-assembled genomes across all sites. CO-oxidizing microbes were present across soil ages, with detectable activity in the younger soils and greatest activity in the oldest, suggesting that these microbes actively use CO as an energy source even in soils with primary vegetation, contrary to general understanding. The findings highlight the intricate dynamics of microbial succession in volcanic soils and challenge conventional expectations about community complexity over time. Understanding pioneer communities elucidates soil restoration processes, which will become critical when countering anthropogenic soil degradation.

microbiology↗

bioRxiv2025

Cross-Fostering with control dams rescues Gut Dysbiosis and Chromatin-associated Transcriptional Changes in Offspring of Opioid-Exposed Dams

Prenatal opioid exposure disrupts gut homeostasis and causes gastrointestinal complications in offspring, but the mechanisms remain unclear. Here using a murine model of prenatal hydromorphone exposure, we examined gut microbiota, intestinal injury, transcriptomic signatures, and chromatin accessibility. Exposed pups displayed marked dysbiosis, epithelial damage, and upregulation of inflammatory gene programs accompanied by relaxed ileal chromatin. Cross-fostering to opioid-naive dams restored microbial diversity, reestablished metabolite-producing taxa, and reversed injury-associated transcriptional and chromatin changes. Fecal microbiota transplantation from exposed dams recapitulated intestinal injury, indicating a microbiome-driven mechanism. These findings reveal a novel gut-microbiome-epigenome axis underlying opioid-induced injury and highlight early microbial intervention as a potential strategy to mitigate developmental harm.

microbiology↗

bioRxiv2025

Mapping Allosteric Rewiring in Viral RNA: Sequence-Encoded Control of Protein Binding Mechanisms

RNA recognition by proteins is governed not only by static structure but also by allostery encoded within non-local dynamic motifs. In this study, we systematically identify allosteric communication hubs in RNA and map multiple residue-connected pathways, revealing how these networks are rewired upon mutation and protein binding. To capture these effects under physiological salt conditions, we performed tens of microseconds of atomistic and steered molecular dynamics simulations and computed binding free energies for Tat-TAR complexes across three immunodeficiency virus variants, BIV, HIV-1, and HIV-2. Allosterically coupled sites were identified using contact-based principal component analysis, and communication pathways were traced through an extended graph-network algorithm--the first such application to RNA systems. Two distant motifs--the bulge and the apical loop--emerge as allosteric switches and information hubs: the bulge engages Tat, while the loop interacts with another protein partner, CycT1, both essential for transcriptional activation and antiviral targeting. We find that HIV-2 TAR, with strong loop-bulge coupling and high self-integrity, favour conformational selection and exhibits lower Tat-binding affinity. In contrast, a single C24 insertion in HIV-1 TAR reconfigures communication pathways, enabling an induced-fit mechanism with enhanced affinity. The study not only elucidates an allosteric rewiring between the loop and bulge but also highlights how this communication is dynamically reconfigured upon protein binding. Tat association at the bulge reorganizes and reorients loop residues, thereby promoting the subsequent recruitment of CycT1. This work overall underscores how sequence (even a single mutation) encoded RNA allostery can modulate not only a proteins binding mechanism and affinity but also influence downstream molecular events within transcriptional signalling cascades. STATEMENT OF SIGNIFICANCEIn vitro selection strategies such as SELEX have long underscored the therapeutic relevance of the TAR RNA-Tat protein interface, yet they remain experimentally demanding and often overlook how subtle sequence variations reshape molecular recognition and RNA-protein binding affinity. By integrating atomistic and free-energy simulations with a graph-network-based framework, the study reveals that even a single-nucleotide change can rewire RNAs allosteric communication pathways, switching between conformational selection and induced-fit mechanisms and alter binding affinity. These findings bridge molecular dynamics with experimental selection, offering a predictive, thermodynamics-based framework to rationalize sequence-structure-function relationships and guide the rational design of high-affinity RNA-protein complexes and next-generation RNA therapeutics.

biophysics↗

bioRxiv2025

Development and validation of an ultra-low-cost, open source normothermic ex vivo organ perfusion platform

BackgroundNormothermic ex vivo organ perfusion (NEVOP) promises to catalyze organ preservation, therapeutic discovery, and organ-specific disease modeling. Existing technology platforms remain inaccessible for research due to restricted access to commercial organ perfusion devices, high costs of both devices and proprietary consumables, and steep technical learning curves. Additionally, the available technology is not optimized to perfuse smaller organs such as the kidney. MethodsTo overcome these barriers, a custom NEVOP circuit was developed using recycled, repurposed, and low-cost components. Porcine kidneys and autologous blood were used to iteratively optimize circuit design. A porcine kidney autotransplantation protocol was adapted to evaluate in vivo kidney function after ex vivo perfusion. To pilot the flexibility of this system as a multi-organ platform for ex vivo human biology, non-transplantable human donor kidney, spleen, and pancreas specimens were stably perfused using human blood products and analyzed. ResultsAn ultra low-cost NEVOP system engineered to perfuse porcine kidneys and diverse human organs (kidney, pancreas, and spleen) sustained viable organs for up to 24 hours with evidence of both function and viability. Key innovations included a parallel flow resistor to facilitate low-flow perfusion in non-heparinized organs and a containment bag with adjustable magnets to provide vascular stability and recycling of venous overflow. The circuit costs less than 1,500USD to construct, and porcine kidneys perfused for 24 hours on this platform demonstrated healthy in vivo function upon autotransplantation. ConclusionsCustom NEVOP platforms constitute novel and potentially transformative research platforms which use low-cost and readily available materials. Paired with access to non-transplantable research organs from altruistic donors, this model provides a road map for investigators to advance biomedical discovery and human ex vivo biology.

bioengineering↗